Time-lapse imaging of cell cycle dynamics during development in living cardiomyocyte
| Autor: | Shugo Tohyama, Keiichi Fukuda, Shinsuke Yuasa, Asako Sakaue-Sawano, Hisayuki Hashimoto, Hidenori Tabata, Kazunori Nakajima, Naoto Muraoka, Shinichiro Okata, Nozomi Hayashiji, Kojiro Yae, Tomohisa Seki, Fumiyuki Hattori, Takahiko Nishiyama, Atsushi Miyawaki, Toru Egashira |
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| Rok vydání: | 2014 |
| Předmět: |
Genetically modified mouse
Embryonic Development Mice Transgenic Time-Lapse Imaging Tissue Culture Techniques Mice Ubiquitin Pregnancy In vivo Animals Myocytes Cardiac Molecular Biology Cell Proliferation Fluorescent Dyes biology Cell Cycle Ubiquitination Cell cycle Embryo Mammalian Cell biology Mice Inbred C57BL Microscopy Fluorescence Cell culture biology.protein Immunohistochemistry Female Cardiology and Cardiovascular Medicine Ex vivo |
| Zdroj: | Journal of Molecular and Cellular Cardiology. 72:241-249 |
| ISSN: | 0022-2828 |
| Popis: | Mammalian cardiomyocytes withdraw from the cell cycle shortly after birth, although it remains unclear how cardiomyocyte cell cycles behave during development. Compared to conventional immunohistochemistry in static observation, time-lapse imaging can reveal comprehensive data in hard-to-understand biological phenomenon. However, there are no reports of an established protocol of successful time-lapse imaging in mammalian heart. Thus, it is valuable to establish a time-lapse imaging system to enable the observation of cell cycle dynamics in living murine cardiomyocytes. This study sought to establish time-lapse imaging of murine heart to study cardiomyocyte cell cycle behavior. The Fucci (fluorescent ubiquitination-based cell cycle indicator) system can effectively label individual G1, S/G2/M, and G1/S-transition phase nuclei red, green and yellow, respectively, in living mammalian cells, and could therefore be useful to visualize the real-time cell cycle transitions in living murine heart. To establish a similar system for time-lapse imaging of murine heart, we first developed an ex vivo culture system, with the culture conditions determined in terms of sample state, serum concentration, and oxygen concentration. The optimal condition (slice culture, oxygen concentration 20%, serum concentration 10%) successfully mimicked physiological cardiomyocyte proliferation in vivo. Time-lapse imaging of cardiac slices from E11.5, E14.5, E18.5, and P1 Fucci-expressing transgenic mice revealed an elongated S/G2/M phase in cardiomyocytes during development. Our time-lapse imaging of murine heart revealed a gradual elongation of the S/G2/M phase during development in living cardiomyocytes. |
| Databáze: | OpenAIRE |
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